RNA Levels Research Articles

Upregulation and epigenetic modification of the creatine transporter SLC6A8 in non-small cell lung cancer.

Lung cancer is a major cause of cancer-related death worldwide and effective therapies, besides surgery, are available only for a small proportion of patients. Since cellular respiration is known to be broadly altered in malignant tumors, the cellular processes of respiration can be a potential therapeutic target. One important element of cellular respiration is creatine and its transport by the creatine transporter SLC6A8. Here we describe the expression of SLC6A8 at the RNA and protein level, epigenetic modifications as well as survival analysis in NSCLC tissues and matched controls. We analyzed epigenetic modifications of the SLC68A gene in 32 patients, of which 18 were additionally analyzed by transcriptome analysis. The expression of SLC6A8 at the protein level was assessed by immunohistochemistry using an independent cohort and correlated with clinicopathological data including survival. Kaplan-Meier analysis was performed to analyze the possible effects of the transcriptional levels of SLC6A8 in another separate cohort (n=1925). SLC6A8 loci are epigenetically modified in NSCLC compared with tumor-free controls. SLC6A8 is upregulated in NSCLC at the RNA and protein level. High mRNA expression of SLC6A8 was associated with an overall poor prognosis in lung adenocarcinoma patients and displayed the strongest adverse prognostic effect in male smokers with adenocarcinomas. Results of transcriptome analysis were partially confirmed at the protein level. Our results suggest an important role of creatine and its transport via SLC6A8 in NSCLC.

Effect of microRNA-141-3p, E2F3, CDK3, and KAT2B overexpression on histologic tumor grade and metastasis status in untreated breast cancer tissues

Introduction: Increasing evidence has reported gene expression alterations in breast cancer (BC) tissues, necessitating their investigating to highlight the molecular basis of the disease development or progression. This study investigated the expression of miR-141, E2F3, CDK3, TP53, and KAT2B, and their association with histologic grade and metastasis in BC tissues. Methods: The RNA expression level of miR-141, E2F3, CDK3, TP53, and KAT2B genes was analyzed in 23 BC and 23 normal tissue samples by RT-qPCR. The associations of the expression level of these genes with clinicopathological features of the BC tissue samples were evaluated. The study also explored the correlation between RNA levels of genes and miR-141. Results: Expression of miR-141, E2F3, CDK3, and KAT2B demonstrated significantly higher levels in BC tumor than normal tissues. TP53 expression showed an increase in tumor compared to normal tissues, although it was insignificant. Moreover, increased RNA expression of miR-141, E2F3, CDK3, and KAT2B corresponded to the advanced stage and regional metastasis of BC. Additionally, the results demonstrated a significant correlation between RNA expression levels of miR-141 with CDK3 and E2F3 with KAT2B. Conclusion: Our findings highlighted clinicopathologic indicators that were relevant to aggressive BC. Besides, Correlations between overexpression of miR-141, E2F3, CDK3, and KAT2B in BC tissues suggest regulatory effects. Taken together, it seems results of this study could provide evidence that dysregulation of gene expression contributes significantly to unveiling the underlying molecular basis of BC.

UBQLN1 links proteostasis and mitochondria function to telomere maintenance in human embryonic stem cells

BackgroundTelomeres consist of repetitive DNA sequences at the chromosome ends to protect chromosomal stability, and primarily maintained by telomerase or occasionally by alternative telomere lengthening of telomeres (ALT) through recombination-based mechanisms. Additional mechanisms that may regulate telomere maintenance remain to be explored. Simultaneous measurement of telomere length and transcriptome in the same human embryonic stem cell (hESC) revealed that mRNA expression levels of UBQLN1 exhibit linear relationship with telomere length.MethodsIn this study, we first generated UBQLN1-deficient hESCs and compared with the wild-type (WT) hESCs the telomere length and molecular change at RNA and protein level by RNA-seq and proteomics. Then we identified the potential interacting proteins with UBQLN1 using immunoprecipitation-mass spectrometry (IP-MS). Furthermore, the potential mechanisms underlying the shortened telomeres in UBQLN1-deficient hESCs were analyzed.ResultsWe show that Ubiquilin1 (UBQLN1) is critical for telomere maintenance in human embryonic stem cells (hESCs) via promoting mitochondrial function. UBQLN1 deficiency leads to oxidative stress, loss of proteostasis, mitochondria dysfunction, DNA damage, and telomere attrition. Reducing oxidative damage and promoting mitochondria function by culture under hypoxia condition or supplementation with N-acetylcysteine partly attenuate the telomere attrition induced by UBQLN1 deficiency. Moreover, UBQLN1 deficiency/telomere shortening downregulates genes for neuro-ectoderm lineage differentiation.ConclusionsAltogether, UBQLN1 functions to scavenge ubiquitinated proteins, preventing their overloading mitochondria and elevated mitophagy. UBQLN1 maintains mitochondria and telomeres by regulating proteostasis and plays critical role in neuro-ectoderm differentiation.

Abstract PO-023: CD70 deregulation in follicular lymphoma at diagnosis is associated with relapse and opens new avenues for dual CD19-CD70 CAR-T therapy

Abstract Follicular lymphoma (FL) is the most common indolent B-cell lymphoma. Despite most patients achieve a complete response with first-line treatment, an important fraction of them relapse, which may worsen the prognosis and increase the risk of a histological transformation to aggressive lymphoma. Therefore, there is a need to better stratify the patients at the diagnosis and develop tailored therapies for high-risk patients. Here we have assessed the immune profile (Nanostring® nCounter) in FFPE-derived biopsies in a series of FL patients at diagnosis, homogenously treated with immunochemotherapy regimen (mostly R-CHOP), and followed-up at Hospital Clínic de Barcelona for more than 12 years. By comparing patients who experienced relapse (n=20) with those who did not relapse at any time (NR, n=12), we have found 31 differentially expressed genes, 25 of them up-regulated in the relapse group. Among them, we have further focused on the role of CD70 in FL pathogenesis. mRNA CD70 was higher in those patients who eventually relapsed and correlated with inferior PFS. Moreover, CD70high FL cases bear a limited number of recurrent somatic mutations. We next sought to determine the specific subpopulations expressing CD70 by multiplex immunofluorescence (Vectra Polaris). Interestingly, we have demonstrated it is mainly expressed in tumor cells correlating with inferior PFS. Moreover, a fraction of T cells from the tumor microenvironment also expresses CD70. Intriguingly, higher levels of CD70 were detected in both CD4+ and CD8+ T cells in the relapse group, including T regulatory and T follicular helper cells. CD70 interacts with CD27, its only known ligand, and can be mutually activated. While CD27 RNA levels did not change between groups, CD27 protein was increased in the B cell population in patients who eventually relapse and in TFH cells, while it was down-regulated in CD8+ and CD4+ T helper non-follicular cells. Furthermore, we found an association between the percentage of CD70+ cells within B cell population and CD70+ in CD4+ and CD8+ cells (R=0.51, p < 0.05). To investigate the role of CD70 in FL pathogenesis we generated two CD70 KO cell lines and FL patient derived cells (n=4) using CRISPR/Cas9 technique. By co-culturing T cells from healthy donors with CD70+ or CD70- FL cell lines, we observed CD70+ tumor cells promote CD70 expression in T cells. In primary samples, we have demonstrated that CD70-KO B cells exhibit reduced response to proliferative stimuli. Moreover, RNAseq analysis followed by GSEA pathway assignment indicated that CD70 expression was related to increased angiogenesis, glycolysis and hypoxia. Finally, to advance towards personalized therapies for this high-risk patient and taking advantage of our institution CAR-T program, we have generated a dual CD19-CD70 CAR-T combining an approved academic product, ARI-0001, and a truncated CD27 protein. Finally, we have demonstrated that this novel CAR-T is effective in both cell lines and ex vivo FL spheroids (PDLS, Dobaño-López, Blood Cancer J 2024), while in vivo studies are ongoing. Citation Format: Ferran Araujo-Ayala, Mireia Bachiller, Raluca Alexandru, Pablo Mozas, Salut Colell, María Villalba-Esparza, Judith Mateos-Jaimez, Ferran Nadeu, Andrea Rivero, Maria Ros, Cèlia Dobaño-López, Juan Garcia Valero, Neus Serrat, Marta Gimenez-Alejandre, Ariadna Giro, Daniel Hodson, Armando López-Guillermo, Elias Campo, Dolors Colomer, Alba Maiques-Diaz, José Ignacio Martín-Subero, Sílvia Beà, Laura Magnano, Sonia Guedan, Carlos E. de Andrea, Patricia Pérez-Galan. CD70 deregulation in follicular lymphoma at diagnosis is associated with relapse and opens new avenues for dual CD19-CD70 CAR-T therapy [abstract]. In: Proceedings of the Fourth AACR International Meeting on Advances in Malignant Lymphoma: Maximizing the Basic-Translational Interface for Clinical Application; 2024 Jun 19-22; Philadelphia, PA. Philadelphia (PA): AACR; Blood Cancer Discov 2024;5(3_Suppl):Abstract nr PO-023.

High Fatty Acid-Binding Protein 4 Expression Associated with Favorable Clinical Characteristics and Prognosis in Papillary Thyroid Carcinoma.

Fatty acid-binding protein 4 (FABP4), a fatty acid transporter that coordinates lipid metabolism, is reported to exert a tumorigenic role in certain cancers. We investigated the effects of FABP4 in the carcinogenesis of thyroid cancer. Bioinformatics data about FABP4 in thyroid cancer were collected from the Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA). Sixteen paired papillary thyroid cancer (PTC) tissues from Taipei Medical University (TMU) were gathered, and commercial thyroid cancer complementary (c)DNA and tissue arrays were purchased to measure FABP4 messenger (m)RNA and protein levels. By analyzing data from the GEO and TCGA, we showed that FABP4 mRNA was reduced in PTC and follicular thyroid carcinoma (FTC). In addition, a lower FABP4 mRNA level in PTC was associated with poor clinical parameters and outcomes in the TCGA database. Moreover, FABP4 transcripts and proteins were downregulated in PTC and FTC, and its mRNA expression was associated with PTC staging in clinical specimens. In the TCGA database and TMU cohort, FABP4 mRNA levels were associated with thyroglobulin (r = 0.511 and r = 0.656, respectively), thyroid peroxidase (r = 0.612 and r = 0.909, respectively), and sodium iodide symporter (r = 0.485 and r = 0.637, respectively) transcripts. In conclusion, FABP4 mRNA and protein levels were reduced in PTC and FTC, and may be used as a potential indicator for thyroid cancer evolution in clinical settings. Further, well-designed research to dissect the molecular mechanism of FABP4 in modulating thyroid carcinogenesis is needed.

Molecular transmission network and drug resistance in treatment-naïve HIV-1 infected patients in the Liangshan District, China.

Objective This study aimed to investigate the molecular transmission network and drug resistance in treatment-naïve HIV-1 infected patients in the Liangshan District, China. Methods The research subjects for this study were HIV-1 infected patients who did not receive any anti-retroviral therapy (ART) in the Liangshan District between January 2022 and July 2023. Peripheral venous whole blood samples were collected from the research subjects. 2 mL blood was used for CD4+ T lymphocyte counting detection. 10 mL blood was centrifuged to separate the plasma and blood cells for quantitative detection of HIV-1 RNA and DNA and drug resistance testing of HIV-1. Results A total of 156 participants were included in this study (88 males and 68 females). The median age of the participants was 37 years old. The findings revealed a positive correlation between the HIV-1 DNA and the HIV-1 RNA levels (r=0.478, P<0.001). However, a negative correlation was observed between HIV-1 DNA levels and CD4+ T lymphocyte counts (r=-0.186, P=0.020). Of the 156 participants, 148 were successfully tested for drug resistance of HIV-1 RNA and HIV-1 DNA simultaneously. Four cases failed the HIV-1 RNA drug resistance testing, and another four failed the HIV-1 DNA drug resistance testing. The most common HIV-1 subtype was the CRF07_BC recombinant. In this study, the overall incidence of transmitted drug resistance (TDR) was 8.33%. The resistance rates of non-nucleoside reverse transcriptase inhibitor (NNRTI) and protease inhibitor (PI) were 7.69% and 0.64%, respectively. Additionally, 32 participants were found to have drug-resistant mutations. The primary drug-resistant mutations were K103N, V179D, E157Q, and A128T, mainly against efavirenz (EFV) and nevirapine (NVP) resistance. Conclusion The drug resistance of HIV-1 infected ART-naïve patients in the Liangshan District cannot be ignored. HIV-1 drug resistance testing is recommended before initiating ART.

Abstract A012: Targeting proteasome vulnerabilities for the treatment of monosomy 7 associated blood disorders

Abstract Monosomy 7 is among the most frequent cytogenetic abnormalities in hematological disorders and is associated with short survival and drug resistance. Despite its high prevalence and detrimental impact, the therapeutic vulnerabilities underlying monosomy 7-associated blood disorders remain largely elusive, impeding progress toward improved patient care. The homeostatic cellular requirement for a normal dosage of essential genes creates an opportunity to target vulnerabilities that arise due to reduced levels of proteins encoded by a haploinsufficient essential gene. Briefly, the loss of one copy of a dosage-sensitive essential gene (gene X), in combination with the inhibition of itself or a related gene (gene Y), or an associated pathway results in lethal consequences for cells. The remarkable and selective clinical efficacy of lenalidomide for the treatment of del(5q) MDS has demonstrated how allelic haploinsufficiency underlies the sensitivity to this drug by synthetic lethality. Differential expression analysis of gene and protein expression in primary AML samples with monosomy 7 revealed significant downregulation of multiple proteasome pathway members at the protein level, but not at the RNA level. Primary AML samples with -7/del(7q) exhibited increased sensitivity (low IC50) to the proteasome inhibitor bortezomib, as evidenced by two independent ex vivo drug screening cohorts (the Beat AML and the FIMM study). Chromosome 7 harbors four proteasome subunits, PSMA2, PSMC2, PSMG3, and SEM1. We performed gene expression, protein expression, copy number analysis, and individual gene knockout experiments. The results have revealed PSMA2 to be a haploinsufficient essential gene on chromosome 7. PSMA2 knockout confers leukemia a growth disadvantage for multiple AML cell clines in both TP53 wild-type and knockout backgrounds. We generated PSMA2 isogenic hemizygous deletion and diploid single-cell clones. PSMA2 hemizygous deletion cells exhibited approximately half the protein expression compared to diploid controls, confirming that PSMA2 is a haploinsufficient gene. PSMA2hemizygous deletion single-cell clones showed significantly enhanced sensitivity to all three evaluated proteasome inhibitors (bortezomib, ixazomib, and carfilzomib), aligning with the sensitivity observed in primary -7/del(7q) leukemia samples. PSMA2 hemizygous deletion cell clones displayed increased p38 and decreased pERK levels upon treatment with proteasome inhibitors, potentially contributing to their increased sensitivity to proteasome inhibitors. Proteomics analysis and in vivo validation is ongoing. As such, we have identified haploinsufficient essential gene PSMA2 mediated proteasome pathway vulnerability in monosomy 7 associated leukemia and further showed that proteasome inhibitors as promising therapeutic approaches for treating hematological disorders associated with monosomy 7. Citation Format: Haijiao Zhang, Basil Allen, Daniel Bottomly, Peter Ryabinin, Schannon K. Targeting proteasome vulnerabilities for the treatment of monosomy 7 associated blood disorders [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Expanding and Translating Cancer Synthetic Vulnerabilities; 2024 Jun 10-13; Montreal, Quebec, Canada. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(6 Suppl):Abstract nr A012.

Early antiviral CD4 and CD8 T cell responses and antibodies are associated with upper respiratory tract clearance of SARS-CoV-2.

T cells are involved in protective immunity against numerous viral infections1. Data regarding functional roles of human T cells in SARS-CoV-2 (SARS2) viral clearance in primary COVID-19 are limited. To address this knowledge gap, samples were assessed for associations between SARS2 upper respiratory tract viral RNA levels and early virus-specific adaptive immune responses, for 95 unvaccinated clinical trial participants with acute primary COVID-19. Functionality and quantity of acute SARS2-specific CD4 and CD8 T cell responses were evaluated, in addition to antibody responses. Most individuals with acute COVID-19 developed SARS2-specific T cell responses within 6 days of symptom onset. Early CD4 T cell and CD8 T cell responses were strongly associated with reduced upper respiratory tract SARS2 viral RNA, independent of neutralizing antibody titers. Overall, these findings indicate a protective role for SARS2-specific T cells during acute COVID-19.

Relationship Between Serum HBV-RNA Levels, Sero-biochemical Indices, and Clinicopathological Parameters in Patients with Hepatitis B Virus

Background: Studies have indicated that HBV RNA exhibits a distinct profile, and the plasma amino acid profile significantly correlates with the different stages of HBV infection. Objectives: This study investigates the relationship between HBV RNA and the plasma amino acid profile and the levels of HBeAg, HBsAg, HBV DNA levels, and other clinical indexes in diagnosing chronic HBV infection diseases. Methods: In this observational study, a total of 155 patients were included. They were categorized as Hepatitis B virus carriers, chronic Hepatitis B (CHB), Hepatitis B-related cirrhosis, and hepatocellular carcinoma patients. Following routine laboratory techniques, the DNA, RNA, serological indexes (HBsAg, HBsAb, HBeAg, HBeAb, and HBcAb), biochemical indexes (ALT, AST), and the concentration of amino acids were determined from the serum. Results: Any relationship between different parameters considered was determined with the chi-square test, correlation, and multivariate logistic regression analysis. We observed that the HBV DNA and RNA levels were significantly higher in patients with chronic Hepatitis B compared to other groups (P &lt; 0.01). The HBV RNA levels significantly correlated with methionine levels (P &lt; 0.05) and not with other amino acids considered in this study. Further, the HBV DNA levels emerged as an independent risk factor for HBV RNA levels, and the area under the ROC curve was 0.840 (P &lt; 0.01) when the levels of HBV RNA, HBV DNA, and methionine were combined to diagnose chronic Hepatitis B. Conclusions: Our study shows that the levels of HBV RNA are correlated with methionine metabolism levels, and this relationship can be used to diagnose and predict the efficacy of treatment for different stages of HepatitisHepatitis B virus infectious diseases.

Bulevirtide Combined with Pegylated Interferon for Chronic Hepatitis D.

In a phase 3 trial, bulevirtide monotherapy led to a virologic response in patients with chronic hepatitis D. Pegylated interferon (peginterferon) alfa-2a is recommended by guidelines as an off-label treatment for this disease. The role of combination therapy with bulevirtide and peginterferon alfa-2a, particularly with regard to finite treatment, is unclear. In this phase 2b, open-label trial, we randomly assigned patients to receive peginterferon alfa-2a alone (180 μg per week) for 48 weeks; bulevirtide at a daily dose of 2 mg or 10 mg plus peginterferon alfa-2a (180 μg per week) for 48 weeks, followed by the same daily dose of bulevirtide for 48 weeks; or bulevirtide at a daily dose of 10 mg alone for 96 weeks. All the patients were followed for 48 weeks after the end of treatment. The primary end point was an undetectable level of hepatitis D virus (HDV) RNA at 24 weeks after the end of treatment. The primary comparison was between the 10-mg bulevirtide plus peginterferon alfa-2a group and the 10-mg bulevirtide monotherapy group. A total of 24 patients received peginterferon alfa-2a alone, 50 received 2 mg and 50 received 10 mg of bulevirtide plus peginterferon alfa-2a, and 50 received 10 mg of bulevirtide monotherapy. At 24 weeks after the end of treatment, HDV RNA was undetectable in 17% of the patients in the peginterferon alfa-2a group, in 32% of those in the 2-mg bulevirtide plus peginterferon alfa-2a group, in 46% of those in the 10-mg bulevirtide plus peginterferon alfa-2a group, and in 12% of those in the 10-mg bulevirtide group. For the primary comparison, the between-group difference was 34 percentage points (95% confidence interval, 15 to 50; P<0.001). At 48 weeks after the end of treatment, HDV RNA was undetectable in 25% of the patients in the peginterferon alfa-2a group, in 26% of those in the 2-mg bulevirtide plus peginterferon alfa-2a group, in 46% of those in the 10-mg bulevirtide plus peginterferon alfa-2a group, and in 12% of those in the 10-mg bulevirtide group. The most frequent adverse events were leukopenia, neutropenia, and thrombocytopenia. The majority of adverse events were of grade 1 or 2 in severity. The combination of 10-mg bulevirtide plus peginterferon alfa-2a was superior to bulevirtide monotherapy with regard to an undetectable HDV RNA level at 24 weeks after the end of treatment. (Funded by Gilead Sciences; MYR 204 ClinicalTrials.gov number, NCT03852433.).

Increased transcriptional elongation andRNA stability of GPCR ligand binding genes unveiled via RNA polymerase II degradation.

RNA polymerase II drives mRNA gene expression, yet our understanding of Pol II degradation is limited. Using auxin-inducible degron, we degraded Pol II's RPB1 subunit, resulting in global repression. Surprisingly, certain genes exhibited increased RNA levels post-degradation. These genes are associated with GPCR ligand binding and are characterized by being less paused and comprising polycomb-bound short genes. RPB1 degradation globally increased KDM6B binding, which was insufficient to explain specific gene activation. In contrast, RPB2 degradation repressed nearly all genes, accompanied by decreased H3K9me3 and SUV39H1 occupancy. We observed a specific increase in serine 2 phosphorylated Pol II and RNA stability for RPB1 degradation-upregulated genes. Additionally, α-amanitin or UV treatment resulted in RPB1 degradation and global gene repression, unveiling subsets of upregulated genes. Our findings highlight the activated transcription elongation and increasedRNA stability of signaling genes as potential mechanisms for mammalian cells to counter RPB1 degradation during stress.

GPX4 mRNA levels in the polymorphonuclear neutrophils are negatively correlated with autoantibody production, disease activity, and lupus alopecia in systemic lupus erythematosus

AbstractBackgroundFerroptosis is a type of regulated necrosis and glutathione peroxidase 4 (GPX4) has been recognized as a key enzyme that protects against ferroptosis. However, the significance of GPX4 in polymorphonuclear neutrophils (PMNs) of systemic lupus erythematosus (SLE) has not been explored. So we examined GPX4 mRNA in PMNs and analyzed its association with serological and clinical features.MethodsA single center research from the Department of Rheumatology and Immunology of the Third Affiliated Hospital, Southern Medical University was conducted between December 2020 and September 2022. Real‐time transcription‐polymerase chain reaction analysis was used to determine the expression of GPX4 mRNA in PMNs from patients suffering from different rheumatoid immune diseases and healthy controls. The associations of GPX4 RNA levels in SLE patients with serological and clinical indicators were assessed by Spearman's correlation analysis. Receiver operating characteristic curve analysis was performed to evaluate the diagnostic value of GXP4 mRNA for SLE.ResultsGPX4 mRNA levels were lower in SLE patients than those in healthy individuals, patients with rheumatoid arthritis and axial spondyloarthritis. GPX4 mRNA levels were negatively correlated with the number of positive autoantibodies (r = −0.3072, p = 0.0300), anti‐dsDNA antibody (r = −0.3654, p = 0.0336), antinucleosome antibody (r = −0.4052, p = 0.0263), erythrocyte sedimentation rate (r = −0.3773, p = 0.0069), C‐reactive protein (r = −0.4037, p = 0.0036), and SLE disease activity index (r = −0.3072, p = 0.0300). GPX4 mRNA levels were downregulated in patients with alopecia compared with patients without alopecia (p &lt; 0.05). The diagnostic capacity of GPX4 mRNA achieved high diagnostic accuracy (area under the curve = 0.848) with sensitivity (78.00%) and specificity (80.95%).ConclusionDownregulated GPX4 mRNA in PMNs of SLE patients is negatively associated with the production of antinuclear antibodies, disease activity, and lupus alopecia, suggesting an important role of ferroptosis in SLE, high diagnostic value of GPX4 mRNA in PMNs, and potential therapies targeting GPX4 for SLE patients, especially lupus alopecia.

Long-term trial of protection provided by adenovirus-vectored vaccine expressing the PPRV H protein

A recombinant, replication-defective, adenovirus-vectored vaccine expressing the H surface glycoprotein of peste des petits ruminants virus (PPRV) has previously been shown to protect goats from challenge with wild-type PPRV at up to 4 months post vaccination. Here, we present the results of a longer-term trial of the protection provided by such a vaccine, challenging animals at 6, 9, 12 and 15 months post vaccination. Vaccinated animals developed high levels of anti-PPRV H protein antibodies, which were virus-neutralising, and the level of these antibodies was maintained for the duration of the trial. The vaccinated animals were largely protected against overt clinical disease from the challenge virus. Although viral genome was intermittently detected in blood samples, nasal and/or ocular swabs of vaccinated goats post challenge, viral RNA levels were significantly lower compared to unvaccinated control animals and vaccinated goats did not appear to excrete live virus. This protection, like the antibody response, was maintained at the same level for at least 15 months after vaccination. In addition, we showed that animals that have been vaccinated with the adenovirus-based vaccine can be revaccinated with the same vaccine after 12 months and showed an increased anti-PPRV antibody response after this boost vaccination. Such vaccines, which provide a DIVA capability, would therefore be suitable for use when the current live attenuated PPRV vaccines are withdrawn at the end of the ongoing global PPR eradication campaign.

Comparative Studies on the Efficacy and Safety of Ainuovirine-Based Versus Efavirenz-Based Antiretroviral Therapy in the Management of Persons Living with HIV: A Real-World Study in Guizhou, China.

In China, non-nucleoside reverse transcriptase inhibitors (NNRTIs) are integral to the antiretroviral therapy (ART) regimen for persons living with HIV (PWH), comprising over 80% of such treatments. To broaden treatment options and improve therapeutic effectiveness, Ainuovirine (ANV), a new NNRTI, was authorized for ART in 2021. Nevertheless, the clinical efficacy of ANV and its impact on blood biochemical markers remain somewhat underexplored. This study seeks to evaluate ANV's clinical performance in ART and its influence on relevant treatment parameters. A retrospective analysis was performed on 208 patients treated with an ANV-based regimen from July 2021 to July 2023, monitoring indicator changes from baseline to week 24. The primary endpoint was the proportion of participants achieving HIV-1 RNA levels of less than 50 copies/mL by week 24. Secondary endpoints involved assessing variations in CD4+ T cell counts and blood biochemical markers from baseline. These outcomes were also compared with data from 241 patients treated with an Efavirenz (EFV)-based regimen in the same time frame. The findings suggest that the ANV-based regimen is as effective as the EFV-based regimen in viral suppression (p > .05) and offers superior improvements in lipid profiles, liver function, and immune system indicators, alongside fewer adverse reactions. These results affirm ANV's efficacy and safety as an antiretroviral therapy option, offering Acquired Immune Deficiency Syndrome patients a wider array of treatment possibilities and the potential for better treatment outcomes.

Dual analysis of IL-8 in non-small cell lung cancer: A clinical perspective on protein and RNA evaluation via imaging mass cytometry.

e15068 Background: The cytokine Interleukin-8 (IL-8) plays a pivotal role in the tumorigenesis and progression of non-small cell lung cancer (NSCLC), influencing both the tumor microenvironment and immune escape mechanisms. Given its significance, accurately assessing IL-8 levels at the protein and RNA levels could provide valuable insights into its prognostic value and inform therapeutic targeting strategies. This study utilizes imaging mass cytometry (IMC) to conduct a comprehensive evaluation of IL-8 expression in NSCLC, integrating both protein presence and gene expression analysis within the spatial context of the tumor microenvironment. Methods: Our approach leverages the capabilities of IMC to simultaneously detect protein and RNA levels of IL-8 in NSCLC tissue samples. For protein analysis, antibodies tagged with heavy metal isotopes specifically target IL-8, allowing for precise quantification and spatial localization within the tumor. Concurrently, RNA detection is achieved through hybridization probes designed for IL-8 mRNA, enabling the direct observation of gene expression patterns. This method provides a unique opportunity to correlate IL-8's protein abundance with its mRNA expression, offering insights into the regulatory mechanisms at play and their impact on the tumor's behavior and patient outcomes. Results: Preliminary data demonstrate a marked heterogeneity in IL-8 distribution at both the protein and RNA levels, with distinct patterns emerging that relate to tumor aggressiveness and immune infiltration. High IL-8 expression, identified through our integrated IMC approach, correlates with poor prognosis and reduced response to standard therapies, underscoring its potential as a biomarker for NSCLC management. Furthermore, the spatial analysis afforded by IMC highlights the microenvironmental niches where IL-8-mediated interactions could drive tumor progression and immune evasion. Conclusions: By employing IMC for the dual analysis of IL-8 protein and RNA in NSCLC, our study provides a comprehensive view of this cytokine's role in lung cancer pathology. The ability to visualize and quantify IL-8 expression within the spatial context of the tumor microenvironment opens new avenues for understanding its biological impact and for developing targeted therapies. Our findings advocate for the inclusion of IL-8 in the biomarker panel for NSCLC, with potential implications for patient stratification and personalized treatment planning. Future research will focus on expanding these observations to a larger cohort and exploring the therapeutic modulation of IL-8 as a strategy for improving NSCLC outcomes.

An eQTL-based approach reveals candidate regulators of LINE-1 RNA levels in lymphoblastoid cells.

Long interspersed element 1 (LINE-1; L1) are a family of transposons that occupy ~17% of the human genome. Though a small number of L1 copies remain capable of autonomous transposition, the overwhelming majority of copies are degenerate and immobile. Nevertheless, both mobile and immobile L1s can exert pleiotropic effects (promoting genome instability, inflammation, or cellular senescence) on their hosts, and L1's contributions to aging and aging diseases is an area of active research. However, because of the cell type-specific nature of transposon control, the catalogue of L1 regulators remains incomplete. Here, we employ an eQTL approach leveraging transcriptomic and genomic data from the GEUVADIS and 1000Genomes projects to computationally identify new candidate regulators of L1 RNA levels in lymphoblastoid cell lines. To cement the role of candidate genes in L1 regulation, we experimentally modulate the levels of top candidates in vitro, including IL16, STARD5, HSD17B12, and RNF5, and assess changes in TE family expression by Gene Set Enrichment Analysis (GSEA). Remarkably, we observe subtle but widespread upregulation of TE family expression following IL16 and STARD5 overexpression. Moreover, a short-term 24-hour exposure to recombinant human IL16 was sufficient to transiently induce subtle, but widespread, upregulation of L1 subfamilies. Finally, we find that many L1 expression-associated genetic variants are co-associated with aging traits across genome-wide association study databases. Our results expand the catalogue of genes implicated in L1 RNA control and further suggest that L1-derived RNA contributes to aging processes. Given the ever-increasing availability of paired genomic and transcriptomic data, we anticipate this new approach to be a starting point for more comprehensive computational scans for regulators of transposon RNA levels.

CREB5 as an essential transcription and tumorigenic regulator in basal-like breast and prostate cancer.

1107 Background: Breast cancers (BC) harbor basal-like transcriptional profiles, in which a large subset are pathologically triple negative breast cancers (TNBC). These are hormone therapy insensitive, have limited targeted therapies, and decreased overall survival (OS). Recent studies determined that prostate cancers (PCs) also exhibit luminal- or basal-like transcriptional subtypes. The 30-40% of basal-like PCs exhibit resistance to hormone therapies and have overall worse clinical outcomes. Here we deployed in silico computational models and tumor phenotype testing in vitro and in vivo. This identified that CREB5, a transcription factor commonly amplified or overexpressed in advanced cancers, regulates basal-like BCs and PCs through stem cell-like genes and properties that drive tumor formation. Methods: Protein and RNA levels were examined in 5 subtypes of BC from the TCGA (n=981). Normalized expression and PAM50 signatures were analyzed using data from Decipher tests of PC patients (n=585). Kaplan-Meier curves were used to compare relapse free survival in PC (n=84) and OS in TNBC (n=248). The ALAN (Algorithm for Linking Activity Networks (1) computational tool was used to determine gene-gene similarities of ~20,000 genes based on ~400,000,000 pairwise comparisons in 208 tumors. In PC (LNCaP) and TNBC (MDA-MB-231 and HCC1806) cell lines, we examined gene expression profiles from RNA-seq and found Hallmark signatures through Gene Set Enrichment Analysis. Motif analysis was conducted from ChIP-seq data on immuno-precipitated CREB5. We conducted in vitro and in vivo assessment of tumorigenicity. Results: Basal BCs harbored increased CREB5 expression and CREB5-high basal BCs had worse outcomes (HR 2.09, 95% CI 1.1-3.96, p&lt;0.021). Reflecting this, in basal-like BC cell lines, CREB5 increased cell viability and formation of tumor-like spheroids in 3D culture. In PCs, CREB5 had a positive correlation with basal- and not luminal-like PCs (Spearman ρ = 0.519, p&lt;0.001). Through ALAN analysis of metastatic PCs, CREB5 exhibited strong concordance with other known resistance genes (FGFR1/2) as well as transcription factors that promote stem cell-like features. Particularly, CREB5 was associated with increased FOSL1 (Spearman ρ = 0.37, p&lt;0.001), a metastasis-regulating gene. In PC cell lines, CREB5 overexpression promoted FOSL1 and other stem cell-like genes and co-bound to FOSL1 transcription regulatory elements. CREB5 overexpression in PC cell lines promoted the formation of tumor-like colonies, spheroids, and increased tumor formation from the xenografts in mice. Conclusions: Basal-like BCs and PCs demonstrate overexpression of CREB5, a gene that promotes tumor formation and stem cell-like genes. This mechanistic knowledge may guide treatment management strategies and aid in the development of novel therapies against basal-like BCs and PCs. 1. Bergom et al, Communications Biology, 2023.

Mechanism study on fig exosome-like nanoparticles inhibiting the progression of bone metastasis in breast cancer.

e12562 Background: Breast cancer has overtaken lung cancer as the most common cancer worldwide. Breast cancer bone metastases can lead to death. How to inhibit the progression of bone metastasis of breast cancer is the key to improve the quality of life of breast cancer patients. In recent years, there have been increasing studies on the therapeutic significance of plant-derived extracellular vesicles (PDEVs), which play an important role in anti-tumor and anti-inflammatory. It is a new potential anti-tumor therapy. According to the Chinese medicine classics, figs have anti-cancer effect, but the main active ingredients are not clear yet. Methods: The exosomes were extracted by differential ultracentrifugation. The breast cancer cells (MDA-MB-231, MCF-7, 4T1, E0771) were treated with 0.1 ug/ml and 0.5 ug/ml fig exosome-like nanoparticles (FELNs) for 48 hours. The proliferation rate was measured by BrdU kit and verified by RT-qPCR. The total RNA from MDA-MB-231 cells treated by FELNs was sequenced to screen the differential genes. Key genes were differentially expressed in MDA-MB-231 and MCF-7 cells and further verified. Animal models of breast cancer in situ and bone metastasis were constructed in mice. FELNs were injected intraperitoneally twice a week, live imaging of small animals was performed on the 14th and 21st day to observe the tumor size and progression of breast cancer, and tibia samples were collected for Trap staining. The bone marrow macrophages of mice were extracted and treated by FELNs. The polarization was detected by RT-qPCR. After the removal of the proteins and nucleic acids in FELNs by protease, DNase and RNase, FELNs intervened in breast cancer cells and macrophages to determine the effective components of the FELNs. Results: After breast cancer cells were treated with FELNs, the proliferation rate was significantly decreased, especially MDA-MB-231 (from 70.3% to 17.7%) and MCF-7 (from 7.32% to 1.63%), and the RNA level of proliferation-related gene were significantly down-regulated. The key gene RN7SL1 was screened by sequencing. Overexpression of RN7SL1 significantly promoted the proliferation of MDA-MB-231 and MCF-7 cells, while the level of proliferation-related gene RNA was significantly up-regulated. FELNs significantly inhibited the tumor size and bone metastasis of breast cancer, and the tibial osteolysis was significantly abated by the treatment of FELNs in tibial Trap staining. After the intervention of FELNs in mouse macrophages, the polarization of M1 macrophages was significantly promoted. The proliferation rate of breast cancer cells increased and the polarization of M1 macrophages disappeared by treatment with protein-free FELNs. Conclusions: The protein components in FELNs inhibit the proliferation of breast cancer cells through RN7SL1, thereby inhibiting the progression of breast cancer bone metastasis.

Cartilage oligomeric matrix protein as a potential biomarker for knee osteoarthritis.

This study aimed to determine the expression and clinical significance of a cartilage protein, cartilage oligomeric matrix protein (COMP), in knee osteoarthritis (OA) patients. A total of 270 knee OA patients and 93 healthy controls were recruited. COMP messenger RNA (mRNA) and protein levels in serum, synovial fluid, synovial tissue, and fibroblast-like synoviocytes (FLSs) of knee OA patients were determined using enzyme-linked immunosorbent assay, real-time polymerase chain reaction, and immunohistochemistry. COMP protein levels were significantly elevated in serum and synovial fluid of knee OA patients, especially those in the advanced stages of the disease. Serum COMP was significantly correlated with radiological severity as well as measures of body composition, physical performance, knee pain, and disability. Receiver operating characteristic curve analysis unveiled a diagnostic value of serum COMP as a biomarker of knee OA (41.64 ng/ml, area under the curve (AUC) = 1.00), with a sensitivity of 99.6% and a specificity of 100.0%. Further analysis uncovered that COMP mRNA expression was markedly upregulated in the inflamed synovium of knee OA, consistent with immunohistochemical staining revealing localization of COMP protein in the lining and sub-lining layers of knee OA inflamed synovium. Most notably, relative COMP mRNA expression in knee OA synovium was positively associated with its protein levels in serum and synovial fluid of knee OA patients. In human knee OA FLSs activated with tumour necrosis factor-alpha, COMP mRNA expression was considerably up-regulated in a time-dependent manner. All results indicate that COMP might serve as a supportive diagnostic marker for knee OA in conjunction with the standard diagnostic methods.

LncTIMP3 promotes osteogenic differentiation of bone marrow mesenchymal stem cells via miR-214/Smad4 axis to relieve postmenopausal osteoporosis.

Promoting the balance between bone formation and bone resorption is the main therapeutic goal for postmenopausal osteoporosis (PMOP), and bone marrow mesenchymal stem cells (BMSCs) osteogenic differentiation plays an important regulatory role in this process. Recently, several long non-coding RNAs (lncRNAs) have been reported to play an important regulatory role in the occurrence and development of OP and participates in a variety of physiological and pathological processes. However, the role of lncRNA tissue inhibitor of metalloproteinases 3 (lncTIMP3) remains to be investigated. The characteristics of BMSCs isolated from the PMOP rat model were verified by flow cytometry assay, alkaline phosphatase (ALP), alizarin red and Oil Red O staining assays. Micro-CT and HE staining assays were performed to examine histological changes of the vertebral trabeculae of the rats. RT-qPCR and western blotting assays were carried out to measure the RNA and protein expression levels. The subcellular location of lncTIMP3 was analyzed by FISH assay. The targeting relationships were verified by luciferase reporter assay and RNA pull-down assay. The trabecular spacing was increased in the PMOP rats, while ALP activity and the expression levels of Runx2, Col1a1 and Ocn were all markedly decreased. Among the RNA sequencing results of the clinical samples, lncTIMP3 was the most downregulated differentially expressed lncRNA, also its level was significantly reduced in the OVX rats. Knockdown of lncTIMP3 inhibited osteogenesis of BMSCs, whereas overexpression of lncTIMP3 exhibited the reverse results. Subsequently, lncTIMP3 was confirmed to be located in the cytoplasm of BMSCs, implying its potential as a competing endogenous RNA for miRNAs. Finally, the negative targeting correlations of miR-214 between lncTIMP3 and Smad4 were elucidated in vitro. lncTIMP3 may delay the progress of PMOP by promoting the activity of BMSC, the level of osteogenic differentiation marker gene and the formation of calcium nodules by acting on the miR-214/Smad4 axis. This finding may offer valuable insights into the possible management of PMOP.